The long-term objectives of this research are to determine the mechanism by which cadmium (Cd) causes bone loss and relate these findings to human exposures. Experiments are designed to test the hypothesis that Cd can act directly on bone cells or on their marrow cell precursors and that these effects of Cd are separate from any effects of Cd on other organs (e.g., renal tubules or gastrointestinal tract). The specific aims of this proposal are to (1) investigate the role of bone cell gene expression in the early bone loss response to Cd in vivo and in vitro; (2) determine if Cd stimulates osteoclast activation by decreasing osteoclast intracellular calcium; (3) further characterize the in vivo acute exposure model for Cd-induced bone loss; and (4) help to link the animal and cell culture results to environmental Cd exposures in humans. Quantitative RT-PCR and transgenic mouse strains will be used to probe for specific genes that might be involved in the bone response to Cd according to current hypotheses. Differential display followed by partial sequencing of the differentially expressed genes will be used to identify genes that might not be expected by current hypotheses. Cd-induced changes in intracellular calcium will be determined in individual osteoclasts cultured on bone wafers using Fura-2 dye and a fluorescence image analysis system. The in vivo acute exposure bone loss model will be characterized with respect to interactions of Cd with ovariectomy and low calcium diet. To ascertain whether environmentally-exposed persons have Cd concentrations in range of those linked to early bone changes, a new Cd assay will be miniaturized and used to determine Cd concentrations in archived blood and urine samples from 188 persons will-characterized with respect to environmental Cd exposures in a zinc smelter town. This will be the first time that samples from persons environmentally exposed to Cd will be evaluated using an assay with a detection limit well below the Cd concentrations in the samples. Animal studies show that bone loss responses occur at blood Cd concentrations in range of levels reported for persons who smoke cigarettes and for workers with low-level Cd exposure in industry. The results suggest that (1) women exposed to Cd are at increased risk of postmenopausal oxteroporosis and (2) the current OSHA action level for Cd in blood may not protect working women from Cd-induced bone loss. The proposed research will address these important issues.